1. Field of the Invention
This invention concerns the field of molecular interactions between two proteins.
More particularly the invention concerns the interactions between an extracellular protein belonging to the thrombospondin family and a receptor, CD47, situated on the surface of the cell membrane.
This invention will have potential applications mainly in the field of tumor, thrombotic and cardiovascular diseases.
More particularly the invention relates to a peptide with the ability to bind specifically the CD47-binding domain of a protein belonging to the thrombospondin family, in such a way as to prevent the binding between the two protagonists.
2. Description of Related Art Including Information Disclosed Under 37 CFR 1.97 and 37 CFR 1.98.
Thrombospondins (TSP), and in particular TSP1 and TSP2, are proteins known to have the ability to bind to the CD47 receptor, which creates a signal inducing a cellular response on the part of the cell. Such an interaction seems to play an important role in particular in fundamental cellular processes, such as the regulation of programmed cell death, also known as apoptosis, or inflammation.
For example, work recently done by the inventors has revealed the anti-apoptotic role of TSP1 on human follicular thyroid carcinoma cells, also known as FTC cells (Rath et al., 2006). Other studies have also shown that a reduction in the expression of TSP1 leads to the reversibility of the phenotype of squamous cell carcinoma thanks to the use of an antisense strategy.
Further research carried out to try and demonstrate the role of TSP1 in the development of breast cancer cells, has, however, had contradictory outcomes; in fact, some of the research has concluded that TSP1 has a pro-apoptotic effect, that is to say that it suggests that it inhibits the growth of the mammary tumor (Esemuede et al., 2004; Manna and Frazier 2004). On the contrary, however, other studies defend the opposite concept, in which TSP1 appears to play an anti-apoptotic role and/or increase the invasive properties of these tumor cells (Wang et al., 1996a, 1996b).
Understanding the molecular mechanisms leading to the interaction between the proteins belonging to the thrombospondin family and their receptors, as well as the discovery of the ensuing cellular responses, therefore constitute an important step in devising therapeutic strategies designed to inhibit the development and proliferation of cancer cells.
Certain prior art documents already recommend using the properties of interaction between thrombospondins and their receptors, in particular CD47, in order to develop strategies either to reduce, or to increase, the rate of apoptosis of cells, whether these are cancerous or not.
Thus, it is known in the prior art, for example U.S. Pat. No. 7,582,725 B2, that an agent binding either the CD47 receptor or thrombospondin-1 can be used to inhibit the binding between said TSP1 and said CD47 receptor. Preventing this interaction is thought to significantly reduce the rate of apoptosis of certain cells such as fibroblasts or epithelial cells, involved in particular in the healing processes, in which they play an important role. Preferentially, with the aim of reducing the rate of apoptosis induced by the TSP1 protein, a peptide with an amino acid sequence with the general structural formula XXYVVM (SEQ ID NO: 4) is used.
However, in the case of this invention, the cells to be targeted and the goal to be reached are completely different. Indeed, the aim in our case is not to try to halt the apoptosis but, on the contrary, to promote the death of the cancer cells (apoptosis and/or necrosis). Thus, the peptide proposed in the aforementioned document is not suitable to solve the problem in question, namely to inhibit the invasive potential of cancer cells, in particular by promoting their entry into apoptosis.
The prior art also includes the use of monoclonal antibodies specifically directed either against the TSP protein, or against the CD47 receptor. This has the consequence of preventing the interaction between these two proteins, and therefore of inhibiting the ensuing cellular responses.
However, the main disadvantage of this technique is that the monoclonal antibodies directed against one or other of the proteins will not specifically bind at the precise site of binding between the TSP and the CD47. Thus, the blocking of the interaction between the two proteins may not be optimal and the fixing of the antibodies, whether on the TSP or on the CD47, may prevent the interaction of these proteins with their other natural ligands. As a result, other cellular processes, which are indispensable in the body and which use one or other of the aforementioned proteins, may also be inhibited or prevented. Furthermore, another major disadvantage of such a strategy resides in the fact that the anti-CD47 antibody may in certain cases act as an agonist of the receptor, that is to say it may activate the latter by means of the interaction.
Patent CA 244 6391 also uses the binding properties between the CD47 receptor and its ligand, the TSP1 protein. More particularly, it is the role of these two molecules in the immune response, and especially in the inflammatory response, which is exploited. The patent thus discloses the use of a monoclonal antibody directed against the CD47 receptor in order, in particular, to inhibit the activity of the suppressor T cells. Such inhibition is thought to play a beneficial role in various cellular processes, in particular in neutralizing infectious agents, allergic reactions, autoimmune or inflammatory diseases, etc.
However, as explained above, the use of a monoclonal antibody directed against one or other of the two proteins does not constitute a satisfactory solution.
Patent WO 2010/017332 A2 also relates to the inhibition of the interaction between the CD47 receptor and thrombospondin. In particular, this patent presents the use of very varied agents to inhibit this interaction prior to the treatment of a patent by radiotherapy, so as to facilitate the surgical removal of the tumor. Indeed, preventing the TSP1/CD47 interaction is thought to favor the protection of the cells in the immune system against the damage caused by their exposure to radiation. As a result, the immune response against tumor cells is thought to be considerably increased.
However, the agents used are not suited to specifically blocking the binding of the TSP to the CD47 receptor.